Many Americans are afflicted by low back pain, degenerative spinal disease, or bone fractures. These musculoskeletal problems are responsible for a major portion of the health care budget and are among the greatest causes of chronic disability and lost productivity in the United States. Orthopaedic surgical treatment of these problems frequently requires bone grafting to promote healing. Fusion of two or more bones with cancellous bone graft may fail to heal in 25-45% of patients, and in even higher percentage of smokers and diabetic patients, co-morbidities which are more prevalent in the veteran population. Use of osteoinductive proteins such as BMP-2 to induce bone formation in these patients is now possible.
In 2002 the U.S. Food and Drug Administration approved rhBMP-2 for use as a bone graft substitute in interbody spine fusions. Despite this regulatory milestone for BMP-2, this technology is not feasible for many patients with bone healing needs due to an unexpectedly high dose required in humans which has resulted in a very high cost (Boden S D, Zdeblick T A, Sandhu H S, and Heim S E. Spine 2000; 25:376-81; Ackerman S J, Mafilios M S, and Polly D W, Jr. Spine 2002; 27:s94-s99). A 15,000-fold higher concentration of BMP-2 is required to induce bone in humans (1.5 mg/ml) than in cell culture (100 ng/ml). Thus, without a dramatic improvement in BMP-2 responsiveness, healthcare economics may severely limit translation of one of the most seminal discoveries related to osteoblast differentiation in the last 50 years from helping large numbers of patients.
Consequently, a further understanding of the complex regulation of BMP-2 during osteoblast differentiation and the cellular responsiveness to such important bone forming proteins is critical so that their effect can be enhanced or their required dose limited to a more affordable quantity of protein especially in the most challenging orthopaedic healing environment—posterolateral lumbar spine fusion.
Several years ago a novel intracellular LIM domain protein critical to fetal and post-natal bone formation was identified (Boden S D, Liu Y, Hair G A et al. Endocrinology 1998; 139:5125-34). Termed LIM Mineralization Protein (LMP-1) it was the first LIM domain protein to be directly associated with osteoblast differentiation. Blocking LMP-1 expression prevents osteoblast differentiation in vitro, suggesting a critical functional role of this novel intracellular protein. Leukocytes expressing the LMP-1 cDNA (via plasmid or adenoviral transduction) that are implanted into rabbits or athymic rats induce bone formation in bony and ectopic locations (Boden S D, Titus L, Hair G et al. Spine 1998; 23:2486-92). The feasibility of LMP-1 delivery by ex vivo gene therapy for spine fusion and bone defect applications in rabbits and primates is currently being evaluated. LMP-1 also has considerable potential as a local, regional, or systemic anabolic strategy for increasing bone density in patients with osteoporosis. However, before clinical applications can be seriously considered it will be critical to understand the mode of action of this protein.
Currently, three splice variants of LIM protein have been identified. These are termed LMP-1, LMP-2, and LMP-3. Human LMP-2 has a 119-base pair (bp) deletion between bp 325 and 444 and a 17-bp insertion at bp 444. The resulting derived protein contains 423 AA with the LIM domains intact and does not induce bone formation when overexpressed in ROB cultures. Human LMP-3 has the same 17 nucleotide insertion at bp 444, resulting in a shift in the reading frame that causes a stop codon to occur at bp 505-507. The resulting 153 AA protein does not have the LIM domains, but overexpression of LMP-3 induces bone formation in osteoblast cultures. Liu et al., J Bone Miner Res., 17(3):406-14 (2002).
It was found that the LMP-1 and the LMP-3 proteins, but not the LMP-2 protein are capable of osteoinduction. The LMP-1 and the LMP-3 proteins possess a relatively short sequence comprising 45 amino acids, which is sufficient to exert an osteoinductive effect.
It has recently been described that this 45 amino acid sequence includes a binding site for a WW-2 domain of a Smurf1 protein. The binding between this osteoinductive amino acid sequence and the WW-2 domain of the Smurf1 protein leads to decreased degradation of Smad1 and Smad5 proteins which are known to mediate the osteogenic effect of the BMP-2 protein. However, more research is needed to provide a better understanding of the role of the LMP proteins in the osteogenesis, as well as the mechanisms by which the LMP proteins regulate osteogenesis. The better understanding of the LMP role may lead to more efficient and affordable therapies of bone defects.